The lymphatic vasculature regulates fluid homeostasis, absorption of dietary lipids and immune system function. The absence of lymphatic vessels is lethal, and impaired lymphatic vessel development causes fluid and protein accumulation in the interstitium, resulting in lymphedema and a persistent inflammatory response. Lymphedema and chronic inflammation are aggravating factors in cardiovascular disease, and are progressive and lifelong conditions affecting millions of people for which curative treatments are not available. Therapeutic agents that can promote lymphangiogenesis should prevent lymphedema and benefit individuals with cardiovascular pathologies. Generation of such agents requires understanding of the cellular and molecular mediators of lymphangiogenesis. The lymphatic vasculature forms a stereotyped, hierarchical branching network that covers the skin and most internal organs of the body. Lymphatic vessels grow by sprouting, and subsequently mature into collecting vessels containing valves that ensure fluid transport. How lymphatic endothelial cells (LECs) interact during sprouting to form a stereotyped network, and how their maturation is regulated remains largely unknown. We will investigate the role of Neuropilin 1 and 2 (Nrp1, 2), co-receptors for vascular endothelial growth factors (VEGFs) and class III Semaphorins (Sema3), in lymphangiogenesis. We have shown that binding of the major lymphangiogenic growth factor VEGF-C to a Nrp2-VEGFR3 complex is critical for LEC sprouting. In addition to VEGF-C-Nrp2-VEGFR3-induced LEC sprouting, our preliminary results show that Sema3A signaling via the other Nrp receptor Nrp1 plays a selective role in lymphatic vessel maturation. We propose to investigate these essential, non-redundant functions of VEGF-C and Sema3A via Nrp2 and 1 in LEC sprouting and maturation, respectively. VEGF-C and Sema3A might be useful to stimulate lymphangiogenesis and prevent lymphedema via distinct cellular and molecular mechanisms. Taken together, this is a comprehensive program aimed at further our understanding of a key biological process, lymphangiogenesis, that may result in development of new therapeutic approaches to prevent lymphedema and cardiac diseases. PUBLIC HEALTH RELEVANCE: The discovery of novel signaling mechanisms controlling lymphangiogenesis will provide new insights and tools into developing new therapeutic strategies for treatment of lymphedema and prevention of atherosclerotic cardiovascular diseases.